Project Summary - Innovation In the first tier of the proposal. Technology Innovation, we are pioneering several technologies for molecular and cell-specific profiling. We are building targeted molecular probes that alter the function of individual proteins or protein complexes, which will be used to map tissue-specific cellular phenotypes. Complex natural product molecules are useful for probing individual proteins and pathways, but they are difficult to chemically synthesize or extract from nature. We are using synthetic DNA to engineer E. coli and yeast hosts for combinatorial biosynthesis of complex natural products. We will use these probes to interrogate human protein complexes using a novel high-throughput protein-protein interaction assay. Probes displaying distinct molecular phenotypes in the protein assay will be analyzed further using a digital microfluidic human cell viability assay. Once the probes are developed and the protein targets are determined, we will assess clinically relevant cell populations. To accomplish this, we will build a platform for the isolation of cancer cells from blood and develop novel methodologies for real-time detection of biomolecules in single cells. These pioneering efforts will shed light on the practical application of these technologies to (1) engineer and utilize natural product probes intractable to chemical synthesis, and (2) enrich rare, living cells for clinical analysis.